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ANALYTICAL CURRENTS Light-emitting nanofibers for lab-on-a-chip applications 500 nm. In both cases, fibers José Moran-Mirabel and col(b) (a) successfully lit up. For fibers leagues at Cornell University Rotating motor spun onto substrates with 500have created light-emitting nm interelectrode gaps, the emisnanofibers by electrospinning. 200 nm sion was easily detected on a The investigators say the fiber Tip scanning CCD camera when voltages as size confines light emission to Substrate HV low as 3.2 V were applied; the nanoscale dimensions, making emission was visible to the naked the fibers attractive for sensing Ground eye at 4 V. and lab-on-a-chip applications. The light from the fibers was Electrospinning produces (a) Schematic of the electrospinning setup to deposit fibers on a micro- and nanofibers from a substrate. Fibers are oriented by the rotating motion of the sub- confined to planar regions of ≤240 325 nm. Moran-Mirabel variety of dissolved materials. strate. (b) Scanning electron micrograph of a 150-nm-diam and colleagues suggest that the The investigators dissolved nanofiber. nanofibers could be easily interuthenium(II) tris(bipyridine) grated as on-chip illumination sources. in dry acetonitrile and mixed in a carrier ters of the fibers ranged from 150 nm The investigators also say different ionic to several micrometers, depending on polymer, poly(ethylene oxide) (PEO). transition metal complexes, with emisthe percentage of PEO in the solution. The blend was electrospun through a sion at other wavelengths, could lead to The grounded silicon substrate was microfabricated tip. A jet of the blend nanofibers capable of locally exciting variformed at the tip, and as the acetonitrile modified with gold interdigitated electrodes. Fibers were spun onto electrodes ous fluorescent tags. (Nano Lett. 2007, evaporated, hardened fibers landed on a 7 , 458– 463) that had interelectrode gaps of 5 µm or grounded silicon substrate. The diame-
Microfluidic system for dynamic microarrays Dynamic microarrays have mobile sub-
the loop and straight channels
strates, usually micrometer-sized beads,
meet and are hydrodynamically
modified with biomolecules or chemicals.
trapped. Once the traps are full,
Shoji Takeuchi and Wei-Heong Tan at the
the flow is redirected to the loop
Institute of Industrial Science (Japan) and
channels so subsequent beads
the Japan Science and Technology Agency
flow out of the device.
have now developed an integrated mi-
focused onto an aluminum pattern
namic microarray. The platform can trans-
located near the trap; localized
port particles, immobilize them for easy
heating results and forms a mi-
signal detection, deliver reagents to them,
crobubble. The expanding bubble
and retrieve selected particles.
displaces the immobilized bead
which are shaped like square waves, superimposed on a straight channel. When
Long dashes
Solid line
For bead retrieval, an IR laser is
crofluidic platform that performs as a dy-
The device has “loop” microchannels,
Dashes
from the trap and into the main
Empty line 100 µm
Beads accumulate to form various types of lines. Arrows indicate the positions of released beads. (Adapted with permission. Copyright 2007 National Academy of Sciences, U.S.A.)
flow. Takeuchi and Tan used the device as a
utes, they introduced 100 beads into the device, perfused analyte over them, and re-
beads first flow into the device, they get
screening tool for the “one-bead–one-com-
trieved the positive beads. (Proc. Natl.
carried into the narrower regions where
pound” combinatorial libraries. In just min-
Acad. Sci. U.S.A. 2007, 104, 1146–1151)
© 2007 AMERICAN CHEMICAL SOCIETY
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ANALYTICAL CURRENTS
COINs for Raman-based tissue analysis Beatrice Knudsen, Selena Chan, and col-
could specifically recognize their respec-
leagues at Intel Corp. and the Fred Hutch-
tive proteins in a mixture. Next, the re-
inson Cancer Research Center have devel-
searchers applied the COIN–antibody con-
oped stabilized, functionalized composite
jugates to detect PSA in formalin-fixed,
organic–inorganic nanoparticles (COINs).
paraffin-embedded, human prostate tissue
The COINs act as optical labels for specific
samples. Formalin-fixed tissue is a difficult
proteins in human tissue.
sample because the molecules are hetero-
A COIN is a cluster of silver nanoparti-
geneously distributed and the tissue auto-
cles that are aggregated in the presence of
fluoresces. However, with the COIN–anti-
Raman-active molecules. Knudsen, Chan,
body conjugates, the investigators could
and colleagues modified their COINs so
identify and quantify the signal in tissue
they could attach antibodies to them. They
areas with high levels of autofluorescence
produced COIN–antibody conjugates that
and create a spatial map of PSA expres-
recognized prostate-specific antigen (PSA)
sion in prostate tissue.
30 µm
nign and cancerous prostate epithelial cells.
COINs in tissue staining, the investigators
PSA expression pattern in tissue; each square represents a single acquisition point. Each point is classified as PSA-positive (red pixels) or PSA-negative (gray pixels) on the basis of the intensity of the measured COIN signal.
The investigators confirmed that each
and cytokeratin-18, a protein found in be-
To illustrate the multiplex capabilities of targeted two types of COINs to PSA. They
PSA spots. This also suggested that steric
protein was recognized by its COIN–anti-
found both types could be detected, and
hindrance of COINs wasn’t a major prob-
body conjugate and that the conjugates
their signals deconvoluted, in almost all
lem. (Nano Lett. 2007, 7 , 351–356)
Scanning outside the NMR tube Bernhard Blümich and co-workers at RWTH Aachen University (Germany) have developed a novel method for shimming ex situ NMR. The approach improves the homogeneity of externally generated magnetic fields and allows high-resolution spectral analysis of arbitrarily large samples. Conventional NMR experiments obtain sharp spectral resolution by placing the sample in the center of a large superconducting magnet for analysis and running currents through shim coils to adjust the internal magnetic field to high homogeneity. Although this configuration works for many experiments, sometimes researchers desire nondestructive analyses of large or valuable objects from which a smaller sample cannot be taken. For this purpose, ex situ NMR is the method of choice. Unfortunately, shimming of the exter2602
nal magnetic fields used in 0.25 ppm ex situ NMR to high homogeneity with shim coils 1 0 –1 is impractical, and thus, the technique loses resolving power. This restriction has been addressed many times with sophisticated – 20 – 40 – 60 60 40 20 0 experimental designs, but (ppm) none so far has matched the resolution of conventional NMR. Ex situ 1H-NMR spectra of water showing spectral resolution Blümich and his colafter magnetic-block shimming (solid line) vs the previous leagues shimmed the state-of-the-art method (dashed line). (Adapted with permismagnetic field of a Usion. Copyright 2007 American Association for the Advanceshaped, single-sided mag- ment of Science.) net with four pairs of compared with the previous state-of-theNdFeB magnetic blocks. The magnetic art ex situ 1H-NMR method, the specfield was adjusted by subtly altering the relative positions of the magnetic blocks. tral resolution of a large water sample Both the S/N and resolution were betimproved 30-fold to 0.25 ppm. (Science ter with this approach. For example, 2007, 315, 1110–1112)
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Assessing phosphopeptide isolation methods Several methods for the isolation of phosphopeptides exist, but how well do they work? To answer this question, Ruedi Aebersold and colleagues at ETH Hönggerberg (Switzerland), the Institute for Systems Biology, and the University of Zurich compared the performances of three popular methods. Although the methods generated reproducible results, they isolated different subsets of the phosphoproteome. The researchers say that no single method, therefore, isolates the entire phosphoproteome. One chemical protocol, phosphoramidate chemistry (PAC), and two affinity-based methods, immobilized metal affinity chromatography (IMAC) and a TiO2-coated resin, were tested. With these approaches, phosphopeptides are retained on a solid-phase support while nonphosphorylated peptides are washed away.
The cytosolic proteome of Drosophila melanogaster Kc167 cells was digested and analyzed with PAC, IMAC, and two variants of the TiO2 procedure. In addition to the typical TiO2 resin with phthalic acid, TiO2 with 2,5-dihydroxybenzoic acid (dhbTiO2) was used; studies suggest that this resin reduces nonspecific binding. By comparing MS1 features among replicates of each approach, Aebersold and co-workers discovered that each method reproducibly isolated a subset of peptides from the complex mixture. However, when features were compared among the approaches, the degree of similarity and overlap was low. Each method, therefore, isolates a different but somewhat overlapping subset of peptides. All of the methods were highly specific, so the reduced overlap was not due to the presence of a constant population of
phosphopeptides isolated with a background of varying nonphosphorylated peptides, say the researchers. When MS2 features were compared among the methods, few phosphorylation sites were common to multiple methods; the exceptions were the TiO2 approaches, which isolated ~95% of the same phosphopeptide sites. The researchers also analyzed the properties of the captured phosphopeptides. They observed that phosphopeptides isolated with PAC typically have one phosphorylation site and are 1000–2500 Da. IMAC produces similar results but tends to isolate more peptides with multiple phosphorylation sites. The TiO2 methods are biased toward acidic peptides with one phosphorylation site. The methods, therefore, capture distinct subsets of the phosphoproteome. (Nat. Methods 2007, doi 10.1038/nmeth1005)
Breath analysis without sample pretreatment The link between exhaled breath and some
nonvolatile compounds, such as urea,
fasting, urea and other protein metabolites
internal biochemical processes has long
amino acids, and glucose.
dominate the breath spectrum, supporting
been known, and researchers have at-
Zenobi and colleagues used EESI Q-
the notion that the body burns protein to
tempted to quantitate these changes with
TOFMS to compare exhaled metabolites
make up for the dearth of carbohydrates.
MS. Now, Renato Zenobi and co-workers
under various physiological conditions. For
They could also track metabolic changes
at ETH Zurich (Switzerland) apply extrac-
example, they found that after overnight
upon drinking beer and smoking a cigarette.
tive ESI quadrupole TOFMS (EESI Q-TOFMS)
The authors note that I
the compounds dissolved in the aqueous
H
microdroplets of breath, eliminating the need for
require modification of Q-TOF
commercial instrumenta-
I
Positive ESI I
100
Ions to MS
to breath analysis. This method directly analyzes
their technique does not
H
MS/MSn
0
m/z
and ease of sampling Commercial Q-TOF interface
a whole new class of
make EESI Q-TOFMS an attractive method for
+HV
pretreatment of sample and allowing analysis of
tion and that the speed
medical breath analysis. Setup for EESI Q-TOFMS. I, desolvation gas inlet, used for sampling of breath; H, heating region in the desolvation gas outlet; HV, high voltage. (Adapted with permission. Copyright 2007 Wiley-VCH.)
(Angew. Chem., Int. Ed. 2007, 46, 580–583)
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ANALYTICAL CURRENTS
Apoptosis probe with a quantifiable readout When cells commit the orderly suicide of
thors hypothesized that
6
apoptosis, researchers in fields from em-
their molecules could
5
bryology to cancer biology want to watch.
detect this shift.
Now, Yves Mely and co-workers at the Université Louis Pasteur (France), Kyiv National
4 3
Their probe, F2N12S,
2
mimics the internal
1
Taras Shevchenko University (Ukraine), and
structure of the cellular
the Palladin Institute of Biochemistry
lipid bilayers and local-
(Ukraine) have developed a new quantita-
izes exclusively to the
tive apoptosis probe based on a 3-hydrox-
cell membrane. Simple fluorescence mea-
yflavone (3HF) derivative.
surements showed that cells undergoing
3HF dyes produce two emission bands,
Normal cells (left) and apoptotic cells treated with actinomycin D (right) stained with F2N12S. Colors correspond to T*/N* ratio. annexin V protein. Because F2N12S does not fluoresce in
apoptosis in the presence of F2N12S have a
water, much of the background typical of
one from the normal (N*) emission and one
much lower N* emission than normal cells,
fluorescent molecules is eliminated without
from the emission of a tautomer (T*) form.
whereas T* emission remains relatively un-
the need for a washing step. But the re-
The authors had previously shown that the
changed between the two groups. When di-
searchers believe the probe’s biggest ad-
intensity ratio between these two emissions
rectly compared in flow cytometry experi-
vantage is its ratiometric readout, which
changed when the molecules were ex-
ments with the most commonly used
frees the signal magnitude from variations
posed to neutral versus negatively charged
apoptosis probe, annexin V–FITC, F2N12S
in probe concentration and instrument sen-
phospholipid bilayers. During apoptosis, the
produced virtually identical results. Yet the
sitivity and allows an absolute measure of
cell shuttles anionic phospholipids into the
new probe does not require the presence
apoptosis over a cellular membrane. (J. Am.
outer leaflet of the membrane, and the au-
2+
of Ca and is much more stable than the
Chem. Soc. 2007, 129 , 2187–2193)
Patterned paper for cheap, portable bioassays We’ve had devices made in glass, silicon, PDMS, and other polymers. Now George Whitesides and colleagues at Harvard University have patterned paper with millimeter-sized channels and used it for bioassays. The investigators say the platform can be useful in the field or as a cheap yet effective alternative to advanced clinical technologies. Whitesides and colleagues patterned the hydrophobic photoresist SU-8 onto hydrophilic chromatography paper. The SU-8 formed the “walls” of channels and spatially controlled the distribution of fluids. Capillary action in the channels transported fluid without pumping. The investigators derivatized the patterned paper for colorimetric glucose and protein assays by embedding the reagents in separate test areas. Then they measured artificial samples in clini2604
cally relevant ranges by dipping the bottom of the derivatized paper in a test solution. Fluid filled the entire pattern of channels in ~1 minute, and the colors developed in 10–11 minutes. The color changes roughly corresponded in intensity to the amount of glucose and protein in the samples, and the paper-based assay was comparable in sensitivity to commercially available dipstick assays. Whitesides and colleagues further adapted the paper platform to measure multiple samples simultaneously. In one trial, a researcher could run 20 different samples within 18 minutes. The investigators also determined that dirt, pollen, and graphite powder at levels much greater than expected in the field didn’t interfere with the assays. (Angew. Chem., Int. Ed. 2007, 46, 1318–1320)
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Chromatography paper patterned with photoresist provides a platform to carry out bioassays for glucose (left) and protein (right). The darker lines indicate SU-8. (Adapted with permission. Copyright 2007 Wiley-VCH.)
news
Proteotypic peptides in the spotlight Some peptides generated by the digestion of a given protein are just detected more often and more reproducibly than others. In back-to-back articles, two research groups describe their work on these proteotypic peptides. Ruedi Aebersold, Bernhard Kuster, and co-workers at the Institute for Systems Biology, Cedars-Sinai Medical Center, the University of California Los Angeles, Cellzome AG (Germany), ETH Zurich, and the University of Zurich developed a method to predict which peptides are proteotypic for a particular protein. In another study, Edward Marcotte and coworkers at the University of Texas used proteotypic peptides to determine absolute quantities of proteins in multidimensional protein identification technology (MudPIT) proteomics experiments. To predict which peptides are proteotypic for a protein, Aebersold, Kuster,
and co-workers studied the physicochemical properties of known proteotypic peptides. Large-scale, well-characterized yeast proteomics data sets that were generated by four common proteomics platforms (1DE/ESI, 1DE/MALDI, MudPIT/ESI, and MudPIT-ICAT) were studied. A total of 494 physicochemical properties, such as charge, likelihood of forming secondary structures, and hydrophobicity, were assessed. The properties that best predicted whether a peptide would be proteotypic varied according to platform. The predictors for yeast also correctly predicted proteotypic peptides for a human data set. The researchers estimate that with their predictors, at least one proteotypic peptide can be identified for most yeast and human proteins. In the other study, Marcotte and coworkers developed a method for largescale absolute protein expression (APEX)
measurements on MudPIT data. No labels and no added standards are necessary. The algorithm estimates the concentration of a protein in a sample on the basis of the number of its peptides that are detected. Factors such as ionization efficiency and amino acid composition are used by APEX to correct for the fact that some of a protein’s peptides are not proteotypic and will not be detected. After the correction is made, the fraction of peptides contributed by a protein to the injected sample pool becomes proportional to the fraction of its peptides that are observed in the MudPIT experiment. The protein abundances calculated with APEX correspond to known abundances over ~2.5 orders of magnitude, and correlate well with data obtained with other approaches. (Nat. Biotechnol. 2007, 25, 117–124)
PEOPLE Sweedler named associate editor Jonathan V. Sweedler
with the neuroscience and bio-
ested in the roles that such neuromodula-
is the newest associate
engineering programs.
tory compounds play in behavior, learning,
editor of Analytical
Sweedler conducts research
and memory.
Chemistry. He was al-
in bioanalytical chemistry and fo-
ready serving as an ac-
cuses on the development of ana-
from the University of California Davis in
tive member of the
lytical methods for assaying com-
1983 and his Ph.D. from the University of
journal’s editorial advi-
plex microenvironments. Those
Arizona in 1989. After a postdoctoral fel-
sory board when his
techniques involve CE, laser-
lowship at Stanford University, he joined
new appointment took
based detectors, MALDI sampling
the University of Illinois faculty in 1991.
effect on March 1.
techniques, nanoliter-volume
Among the prizes that he has been award-
Sweedler earned his B.S. in chemistry
Sweedler is the William H. and Janet Lycan
NMR, and micro- and nanofluidic sam-
ed are the Heinrich-Emanuel Merck Prize
Professor of Chemistry and director of the
pling. His group applies these methods to
(2002), and the American Chemical Society
Roy J. Carver Biotechnology Center at the
studies of the distribution, metabolism, and
Division of Analytical Chemistry’s Award in
University of Illinois at Urbana–Champaign.
dynamic release of neuropeptides and
Chemical Instrumentation (2002) and
In addition, he is a faculty member of the
classical neurotransmitters in a cell-spe-
Arthur Findeis Award for Young Analytical
university’s Beckman Institute for Ad-
cific manner as well as the roles of neuro-
Scientists (1997). The American Associa-
vanced Science and Technology and Insti-
transmitter cotransmission in well-defined
tion for the Advancement of Science elect-
tute for Genomic Biology and is affiliated
neuronal networks. He is especially inter-
ed him a fellow in 2001.
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PEOPLE 2007 ACS national award winners
From left to right: The 2007 ACS national award winners included Michael D. Fayer, Jean H. Futrell, James W. Jorgenson, J. Michael Ramsey, and George M. Whitesides.
Among the honors presented on March 27 at the 223rd American Chemical Society (ACS) National Meeting and Exposition in Chicago were: Michael D. Fayer won the E. Bright Wilson Award in Spectroscopy for his work in nonlinear optical spectroscopy. Fayer is the David Mulvane Ehrsam and Edward Curtis Franklin Professor of Chemistry at Stanford University. The prize recognizes numerous contributions that include the transient grating and photon echo techniques and ultrafast IR vibrational echo experiments. Jean H. Futrell earned the Frank H. Field and Joe L. Franklin Award for Outstanding Achievement in MS, which is sponsored by Waters Corp. Futrell is currently a Battelle Fellow at the Pacific Northwest National Laboratory. His contributions include tandem MS, including the triple quadrupole; mechanisms and dynamics of low-energy ion– molecule reactions; the ion funnel; and the understanding of fundamental ionic phenomena such as collisional activation and energy transfer of ions at surfaces. James W. Jorgenson won the ACS Award in Analytical Chemistry, which is sponsored by the Battelle Memorial Institute. Jorgenson, a former associate editor of Analytical Chemistry, is the William Rand Kenan, Jr., Distinguished Professor of Chemistry at the University of North Carolina at Chapel Hill. The 2606
award recognizes his impact on the field and on society with the development of CE, which made rapid sequencing of the human genome possible, as well as ultrahigh-pressure LC, multidimensional separations, and single-cell analysis. J. Michael Ramsey received the ACS Award in Chromatography, which is sponsored by Supelco, Inc. He is the Goldby Distinguished Professor of Chemistry at the University of North Carolina at Chapel Hill and a former editorial advisory board member of Analytical Chemistry. Ramsey is best known for his work in lab-on-a-chip technology
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and nanofluidics, and he has made contributions in MS. George M. Whitesides received the Priestley Medal, which is sponsored by ACS, and recognizes distinguished service to the chemistry community. He is the Woodford L. and Ann A. Flowers University Professor at Harvard University. His research interests range quite broadly, but he currently emphasizes areas such as physical and organic chemistry, materials science, surface science, microfluidics, self-assembly, micro- and nanotechnology, and science for developing economies.
Analytical chemist wins Grainger Challenge Abul Hussam, a chemistry professor at George Mason University, won the National Academy of Engineering’s Grainger Challenge Gold Award ($1 million) for his household system for removing arsenic from drinking water. Hussam earned his B.S. and M.S. in chemistry at the University of Dhaka (Bangladesh) and his Ph.D. in analytical chemistry from the University of Pittsburgh. The prize was meant to encourage the development and dissemination of technologies to enhance social and environmental sustainability. Water For People won the Silver Award ($200,000), and the Children’s Safe Drinking Water Program at Procter & Gamble Co. earned the Bronze Award ($100,000). In Hussam’s approach, water passes through a bucket filled with river sand, which removes coarse particles, and 20 lb of a composite iron matrix, which removes inorganic arsenic. Then the water flows through a second bucket filled with coarse river sand, wood charcoal (to remove organics), and fine river sand and wet brick chips (to remove fine particles and stabilize the fluid flow).
